The NDP Sensor is a coumarin-labeled form of a bacterial NDP kinase. The sensor autophosphorylates a histidine residue in the presence of NTP (nucleoside triphosphate), producing a highly fluorescent P~NDP sensor (Figure 1). The reverse reaction regenerates the unphosphorylated form. In the presence of excess nucleotides over the NDP Sensor, the ATP/ADP ratio determines the NDP Sensor phosphorylation state via fluorescence intensity.

Table 1. Z´-LYTE™-compatible monochromator-based instruments

Tecan Safire and Safire2™

Excitation filter

400/12*

Emission filter, donor

445/12*

Emission filter, acceptor

520/12*

*Wavelength in nm/bandpass

Figure 1. Mechanism of the NDP Sensor

The NDP Sensor is an excellent universal kinase assay platform for compound profiling and screening (Figure 2), either in endpoint or kinetic mode, and can also be used to identify substrates for uncharacterized kinases (Figure 3).

Table 2. Z´-LYTE™-compatible filter-based instruments

Tecan GENios Pro™

Tecan Ultra™ series

Molecular Devices Analyst®

PerkinElmer EnVision™

Excitation filter

405/20*

405/20*

405/35*

400/25*

Emission filter, donor

465/35*

465/35*

460/40*

460/25*

Emission filter, acceptor

535/20*

535/20*

530/25*

535/25*

Dichroic for excitation

330/420:440/850

50%

425

General dual

Dichroic for emission

330/420:440/850

320/500:520/800

425

General dual

*Wavelength in nm/bandpass

Figure 2. The NDP Sensor as a universal kinase assay The NDP Sensor enables endpoint or real-time kinase assays with peptide, protein, or lipid substrates. All fluorescence measurements were taken at 440/487 nm (Ex/Em) in 384-well plates.

Thermo Fisher Scientific

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